Phomopsis liquidambari is a classical endophytic fungus with great application potential in ecology and agriculture; however, studies on its exopolysaccharides are lacking. Here, we aimed to evaluate the structure and bioactivity of PLN-1, an exopolysaccharide derived from the P. liquidambari NJUSTb1 strain. The structure was elucidated by chromatography/spectral methods and hydrolyzation. Immunomodulation, moisture absorption, and retention properties were investigated after sulfation and carboxymethylation modification. Results showed that PLN-1 contained a linear repeating unit of →[4)- α - d -Glc p -(1→6)- α - d -Glc p -(1→4)- α - d -Glc p -(1→4)- α - d -Glc p -(1→] n , with a molecular weight of 343 kDa. The degrees of substitution of sulfated polysaccharide (S-PLN-1) and carboxymethylated polysaccharide (C-PLN-1) were 1.228 and 0.903, respectively. S-PLN-1 showed stronger moisture absorption and retention properties than PLN (crude EPS), C-PLN1, and PLN-1. Furthermore, PLN, S-PLN-1, and C-PLN-1 stimulated the proliferation of RAW 264.7 cells with no cytotoxicity. The elucidation of PLN-1 in this study paves the way for future applications.
Phenolic acid allelochemicals released by decomposing foliage can have negative impacts on growth of plants and microbes in soil. However, these chemicals can potentially be degraded by broad-spectrum endophytic fungi such as, Phomopsis liquidambari (strain B3), which is capable of growing on phenolic 4-hydroxybenzoic acid (4-HBA) as the sole carbon and energy source. In this study, the optimum concentration of 4-HBA supplied in culture as the carbon source was determined to be 600 mg L −1 , with maximum biomass of fungal growth and, 94% and 99% degraded by P. liquidambari in liquid culture and soil conditions, respectively, within 48 h. The fungal metabolic pathway of 4-HBA was investigated using high performance liquid chromatography–mass spectrometry (HPLC–MS) and gas chromatography–mass spectrometry (GC–MS). The results showed that 4-HBA was first hydroxylated to 3,4-dihydroxybenzoic acid, converted to catechol and then oxidated to cis , cis -muconic acid in the tricarboxylic acid (TCA) cycle. The benzene ring was opened through ortho catechol cleavage. During the course of fungal incubation, activities of three main 4-HBA degradation pathway enzymes exhibited a continuous cascading pattern of induction. 4-HBA hydroxylase showed the highest activity after 8 h of incubation, with 3,4-dihydroxybenzoic acid decarboxylase reaching a peak at 24 h and catechol 1,2-dioxygenase peaking at 32 h, which corresponded with changes in corresponding substrate concentrations. These results indicated the bioremediation potential of this endophytic fungus to degrade accumulated phenolic acid allelochemicals in soil to alleviate the effects of their ecological suppression.
Wang Hong-Wei,Zhang Wei,Su Chun-Lun,Zhu Hong,Dai Chuan-Chao
Phytoestrogens are plant-derived hormonally-active compounds known to cause varied reproductive, immunosuppressive and behavioral effects in vertebrates. In this study, biodegradation of luteolin, a common phytoestrogen, was investigated during incubation with endophytic fungus Phomopsis liquidambari . The optimum concentration of luteolin as sole carbon source supplied in culture was 200 mg L −1 , which allowed 97 and 99 % degradation of luteolin by P. liquidambari in liquid culture and soil conditions, respectively. The investigation of the fungal metabolic pathway showed that luteolin was first decomposed to caffeic acid and phloroglucinol. These intermediate products were degraded to protocatechuic acid and hydroxyquinol, respectively, and then rings were opened by ring-cleavage dioxygenases. Two novel genes encoding the protocatechuate 3,4-dioxygenase and hydroxyquinol 1,2-dioxygenase were successfully cloned. Reverse-transcription quantitative polymerase chain reaction demonstrated that expression levels of mRNA of these two genes increased significantly after P. liquidambari was induced by the intermediate products caffeic acid and phloroglucinol, respectively. These results revealed that P. liquidambari can biodegrade luteolin efficiently and could potentially be used to bioremediate phytoestrogen contamination.
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